The overall goal of our research is to better understand the pathogenesis of Epstein-Barr virus (EBV) infection in order to develop therapies that can reduce, control, or prevent EBV-associated diseases. EBNA-1 is an EBV protein important for the maintenance of the virus episome. EBNA-1 is expressed in all phases of EBV infection, is one of only a few viral genes expressed by infected B cells circulating in the blood of persistently infected hosts, and is the gene most consistently expressed in EBV associated malignancies. EBNA-1 specific cytotoxic T cells are present in persistently EBV infected individuals, but these T cells are unable to effectively kill EBV infected cells due in part to an inhibitory effect of the EBNA-1 glycine-alanine repeat (GAR) domain that prevents appropriate processing and presentation of EBNA-1 peptides to T cells. This application focuses on the importance of EBNA-1 for persistent EBV infection, the importance of EBNA-1 immune evasion for persistent infection, and the possibility of manipulating the EBNA-1 immune response as a therapeutic tool against EBV-associated malignancies. We will use the rhesus lymphocryptovirus (LCV) animal model system for EBV pathogenesis and other laboratory systems to address these issues in the following specific aims: Specific Aim #1. Test whether the ability to downregulate LCV latent gene expression to an EBNA- 1 only promoter is essential for persistent LCV infection. Specific Aim #2. Test whether immune evasion mediated by the EBNA-1 glycine-alanine repeat domain is essential for persistent LCV infection. Specific Aim #3. Use immunosuppression to test whether the EBNA-1 mutant viruses are attenuated due to more effective immune clearance versus a functional defect required for persistent infection. Specific Aim #4. Explore strategies to enhance EBNA-1 recognition and killing of EBV infected B cells by EBNA-1 specific CTLs as a potential therapeutic.